Role of Bailout Gene-Silencing Therapy in Plaque Lipid Reduction: Intravascular Imaging Study

An altered lipid profile, with elevated low-density lipoprotein cholesterol (LDL-C) in the forefront, plays a causal role in the development and progression of atherosclerotic cardiovascular disease [1] and is therefore an essential therapeutic target [2]. The first-line hypolipidaemic pharmacotherapy are statins; however, the maximally tolerated doses may not reduce lipid levels enough to reach the therapeutic targets. Therapy optimization by the addition of nonstatin agents has gained attention, of which ezetimibe is commonly prescribed [3]. However, insufficient effectiveness of treatment with these standard medications is not uncommon. Recently, the role of proprotein convertase subtilisin/kexin type 9 (PCSK9) in LDL metabolism has been emphasized, translating it to the development of novel lipid-lowering agents [4]. PCSK9 pathway plays a crucial role in lipid homeostasis mainly by promoting lysosomal degradation of LDL receptors, resulting in elevated LDL-C plasma levels [5]. Inhibition of PCSK9 is of evolving interest in the context of lipid-lowering pharmacotherapy, and PCSK9-inhibiting monoclonal antibodies are already successfully implemented in clinical practice [6]. A novel medication targeting PCSK9, receiving first approval in 2020, is inclisiran. It is a small interfering ribonucleic acid molecule conjugated to triantennary N-acetylgalactosamine carbohydrates (GalNAc) [7]. The mechanism of action of inclisiran is based on complementary binding of PCSK9 messenger RNA and interference with its translation in hepatocytes, thus reducing the levels of PCSK9 [8]. A significant lipid-lowering effect and sufficient safety profile of the new-generation medication have been established in ORION trials [9, 10]; nevertheless, data concerning practical experience with inclisiran are limited. Intravascular ultrasound (IVUS) and near-infrared spectroscopy (NIRS) allow to assess the dynamics of plaque burden and lipid content in the context of optimal lipid-lowering pharmacotherapy [11, 12]. Intravascular imaging studies demonstrated association of maximal lipid-core burden index within 4 mm (maxLCBI4 mm) of the segment of interest assessed by NIRS with cardiovascular events. Data from LRP study showed non-index culprit-related major adverse cardiovascular event increase in patients having maxLCBI4 mm above 400 [13]. Predictive role of simultaneously increased LCBI and large plaque burden in patients with at least one non-culprit lesion was established in PROSPECT II trial with cut-off values for maxLCBI4 mm of 324.7 and plaque burden of 70% [12]. The aim of this study was to evaluate the impact of optimal hypolipidaemic pharmacotherapy, including add-on inclisiran, on the lipid profile and dynamics of plaque lipid content, evaluated by intravascular imaging, in patients with stable coronary artery disease (CAD).

This was a prospective dual-arm cohort single-centre study that enrolled CAD patients above 18 years of age, with chronic coronary syndrome admitted for elective percutaneous coronary intervention (PCI). After successful PCI, all study participants had undergone intravascular imaging (IVUS and NIRS) of the segment of interest for atherosclerotic plaque assessment (written informed consent for the procedure signed according to regulations in the hospital).

All patients during the rule-in period of 4–6 weeks were on a maximum tolerated dose of statin and/or ezetimibe therapy. In patients having LDL-C level >1.8 mmol/L on maximum tolerated statin and/or ezetimibe therapy, inclisiran therapy was started as add-on treatment in the context of optimal pharmacotherapy (inclisiran group). The first inclisiran subcutaneous injection (284 mg) was administered immediately after the baseline procedure. The second injection was received after 3 months, and every 6 months thereafter, in accordance with the product characteristics document. All injections were administered during ambulatory visits. Patients with LDL-C ≤1.8 mmol/L continued receiving maximally tolerated statin dose and/or ezetimibe (statin/ezetimibe group). High-intensity statin therapy was defined as atorvastatin 40 mg and 80 mg or rosuvastatin 20 mg and 40 mg. Patients admitted due to acute coronary syndrome with severe coronary calcification, chronic heart failure functional class III–IV (NYHA), and stage IV–V chronic kidney disease were considered ineligible.

Baseline data concerning demographics and medical history were registered. Blood test results (lipid profile changes) were evaluated at baseline, during index procedure and after 3, 6, 9, 12 and 15 months in the inclisiran group and after 15 months in the second group. The patients underwent follow-up coronary angiography, IVUS and NIRS investigations that were performed 15 months (±1 month) after the index procedure. Clinical and demographic data were collected from hospital charts, reviewed by qualified personnel blinded to the study objectives, and prospectively entered into a database.

All participants signed informed consent, confirming participation in this study, which was approved by the Ethics Committee of Riga Stradins University (2-PEK-4/242/2022) and complied with the Declaration of Helsinki (WMA, 2013). The trial has been registered in the ClinicalTrials.gov database (NCT05639218).

In the study participants, successful PCI was followed by intravascular imaging. The segment of interest was determined in the proximal or middle third of a coronary artery at least 40 mm in length, with nonobstructive 20–50% atherosclerotic plaque according to the coronary angiography findings. NIRS-IVUS-automated Dualpro^™ catheter (Infraredx, Inc., Burlington, MA, USA) pullback 0.5 mm/s was performed from at least 10 mm distal to the lesion to the coronary ostia. Quantitative IVUS measurements included the cross-sectional areas of the external elastic membrane, lumen, plaque burden, and minimal luminal area. External elastic membrane and luminal borders were contoured for frames at 1-mm intervals in matched regions of the target segments. With NIRS investigation, the total plaque LCBI was evaluated, and maxLCBI4 mm was estimated. Intravascular imaging was performed at 15-month follow-up of identical segments detected with help of anatomical markers (bifurcations, side branches, etc.). The IVUS/NIRS analysis was performed by two independent investigators blinded to clinical and angiographic findings.

Data analysis was carried out with IBM SPSS Statistics 23.0 software. Measures of central tendency for continuous variables were presented as the mean and standard deviation, as well as the median and interquartile range. Statistical significance of baseline and follow-up changes of lipid levels and intravascular imaging-derived parameters among participants in each group was established by non-parametric Wilcoxon test. Between-group differences for values in inclisiran and statin/ezetimibe groups were determined using Mann-Whitney test. Statistical significance was considered for p < 0.05.

The patient baseline characteristics are summarized in Table 1, referring to data before the rule-in period. No significant differences were established between the inclisiran group and the statin/ezetimibe group. The mean baseline LDL-C was 2.88 ± 1.16 mmol/L in the inclisiran group and 2.76 ± 1.28 mmol/L among patients receiving statins and/or ezetimibe only.

Hypolipidaemic therapy changes throughout the study period are demonstrated in Figure 1 as a Sankey diagram. One patient in the inclisiran group had statin intolerance, therefore being only on ezetimibe therapy besides inclisiran injections. In the inclisiran group, approximately 80% of patients received ezetimibe as a part of maximum lipid-lowering treatment.

Changes in lipid profile parameters are represented in Table 2. Baseline values refer to blood test results before rule-in period. Index procedure results were obtained at the moment of initial IVUS and NIRS investigation. LDL-C level changes were estimated referring to the moment of initiation of maximum lipid-lowering therapy – baseline for statin/ezetimibe group and index procedure values for inclisiran group. The follow-up lipid level results are displayed as the mean and the median at 15 months. After 15-month treatment, 12 out of 24 patients receiving inclisiran reached LDL-C target <1.8 mmol/L. Absolute LDL-C changes among patients who received inclisiran and those who took statin/ezetimibe are represented in Figure 2.

The relative LDL-C difference compared to baseline in the patient group receiving inclisiran at the time of 3-, 9-, 12-, and 15-month follow-up in relation to administered inclisiran injections is summarized in Figure 3. When follow-up blood tests were performed soon after inclisiran injection, the lipid-lowering effectiveness was higher compared to blood tests acquired at the time of the next injection after 6 months.

Individual response to inclisiran therapy was evaluated. The interindividual variability of mean relative LDL-C reduction after 15 months of inclisiran treatment is represented in Figure 4.

The intravascular imaging (NIRS and IVUS) results of the segment of interest and the comparison between patients in the inclisiran and statin/ezetimibe groups are summarized in Table 3. No significant procedure-related complications were observed. In 2 patients receiving inclisiran follow-up, intravascular imaging was not performed. LAD was analysed in 45% of cases (n = 18), LCx in 20% (n = 8), and RCA in 35% (n = 14) of study participants.

The main finding in this study was stabilization of atherosclerotic plaque by NIRS-assessed lipid content decrease with add-on inclisiran in patients with insufficient effectiveness of statin and/or ezetimibe therapy similar to plaque composition changes in statin/ezetemibe treatment responder group. In our study, the intravascular imaging results demonstrated a significant reduction in maxLCBI4 mm in the inclisiran and statin/ezetimibe groups when comparing baseline and the 15-month follow-up results, with no statistically significant difference between the groups. Prognostic role of maxLCBI4 mm has been established in randomized clinical trials [11, 12]; thus, lipid-lowering therapy-induced plaque lipid content reduction is presumably a marker of atherosclerotic plaque stabilization. The first published data in the field showed controversial results. The YELLOW trial proved that in obstructive lesions, even short-term intensive statin therapy can reduce plaque lipid content [14], while in IBIS-3 trial there was no effect of long-term high-dose statin on analysed atherosclerotic plaque characteristics, assessed by IVUS and NIRS imaging. In the PACMAN-AMI trial [15], the addition of PCSK9-inhibiting therapy after acute myocardial infarction with the monoclonal antibody alirocumab demonstrated significantly greater LCBI reduction after 52 weeks compared to the effect of high-intensity statin monotherapy. In the recently reported YELLOW III trial results, similar findings regarding LCBI reduction during 26 weeks of evolocumab therapy were detected in stable CAD patients [16]. However, different design of our study does not allow direct comparison of outcomes to abovementioned trials. In our study, inclisiran was started after 4–6 weeks of statin/ezetimibe therapy if LDL-C levels were >1.8 mmol/L. Thus, inclisiran-receiving patients were compared to well-controlled patient cohort having LDL-C <1.8 mmol/L on standard treatment while other PCSK9 inhibitor trials randomized patients that did not achieve LDL-C target [15, 16]. Comparable cholesterol management in both groups explains similar plaque lipid content reduction in our study. Moreover, in our research, for patients not reaching the LDL-C target, inclisiran was added to statins at the maximum tolerated dose, and a high proportion of patients taking ezetimibe were present: 83.33% versus 0.7% in PACMAN-AMI [15]. In the PRECISE-IVUS trial, ezetimibe addition to the therapy demonstrated significantly lower LDL-C levels and greater plaque regression than statins only [17]. Besides quantitative plaque changes, in a single-centre optical coherence tomography study, the statin plus ezetimibe combination showed a greater increase in fibrous cap thickness along with lower LDL-C levels [18].

Analysing plaque parameters, we observed percent atheroma volume decrease in inclisiran-receiving patients. Similar findings were detected in CAD patients treated with alirocumab and evolocumab in PACMAN-AMI [15] and GLAGOV [19] trials, respectively. Nevertheless, in all previously mentioned trials quantitative plaque parameter change was numerically small – decrease by 1.67% in inclisiran group in our study, 0.95% with evolocumab in GLAGOV trial [19], and 2.13% with alirocumab in PACMAN-AMI trial [15]. Thus, LDL-lowering-induced qualitative plaque composition change should have important contribution to clinical benefits in CAD patients. Nevertheless, further research is needed to compare the effects on atherogenesis and plaque stabilization of different PCSK9-reducing mechanisms – circulating PCSK9-inhibiting monoclonal antibodies and PCSK9 gene silencing by inclisiran. Besides, the plaque stabilization response to PCSK9 inhibition could be genetically determined.

The causal role of LDL-C in atherosclerotic cardiovascular disease and plaque progression has been stated in the European Atherosclerosis Society consensus document [20]. In the real-world scenario, long-term adherence to statin therapy is poor [21], and a significant number of patients do not achieve LDL-C targets, remaining at high risk of cardiovascular events [22, 23]. Patients report an inability to tolerate full therapeutic doses of statins with an impact on compliance, highlighting the need for additional treatment [24, 25]. In our study, patients with high LDL-C levels, despite being on maximally tolerated statin dose and/or ezetimibe therapy with mean LDL-C 2.65 ± 1.05 mmol/L, were assigned to receive add-on inclisiran, resulting in significant reductions in the LDL-C levels, which was a decrease of 26.42% for data after 15 months of therapy and 36.17% for pooled follow-up results during 15 months. All inclisiran injections were administered in an ambulatory follow-up setting, thus contributing to solutions for nonadherence problems and improving the overall cardiovascular risk profile. ORION trial programme findings demonstrated time-adjusted LDL-C reductions of 51.3% and 45.8% in the ORION-10 and ORION-11 trials, respectively [10]. The milder effect of inclisiran in our study can be explained by selected drug resistant patient cohort and significantly higher ezetimibe usage compared to ORION trial programme. Moreover, similar LDL-C reduction in both statin/ezetimibe and inclisiran groups is explained by the study design. Inclisiran receiving patient group was compared to patients reaching LDL target within 4–6 weeks of statin/ezetimibe therapy. Additionally, a significant reduction in total cholesterol (TC) by 15.65% and triglycerides (TG) by 29.31% was also noted in our research. By comparison, in the ORION-11 trial at day 510, the TC reduction was 28.0%, and the TG levels were lowered by 12.0%. The ORION trial results and our findings demonstrated interindividual variability and a relationship between LDL-C reduction and the timing of follow-up blood tests [10]. The limitations of our study are the small sample size and lost to follow-up in 2 patients due to refusal of repeated coronary intervention in inclisiran group.

The addition of bailout inclisiran in the context of optimal lipid-lowering pharmacotherapy led to remarkable LDL-C decrease along with plaque stabilization by lipid content reduction. Nevertheless, future studies are needed regarding the effects of PCSK9-inhibiting agents with different mechanisms, including a focus on pathogenetic pathways in atherosclerotic plaques, such as local PCSK9 expression.

This study was reviewed and approved by the Ethics Committee of Riga Stradins University (Approval No. 2-PEK-4/242/2022) and complied with the Declaration of Helsinki (WMA, 2013). All participants signed informed consent, confirming participation in this study.

The authors have no conflicts of interest to declare.

Inclisiran was provided by Novartis company.

K.T.: conceptualization, methodology, investigation, and writing – original draft; B.K. and M.L.: data curation, formal analysis, investigation, and writing – original draft; M.K., E.K., L.C., S.J., I.N., D.S., and I.K.: investigation and writing – review and editing; A.G.: investigation; and A.E.: conceptualization, methodology, investigation, and writing – review and editing.

Data generated or analysed during this study that support the findings are included in this article. Further enquiries can be directed to the corresponding author B.K. upon reasonable request.